MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'Biological Procedures Online' source. Thu, 09 Feb 2012 16:56:37 +0100 http://www.medworm.com/index.php?rid=5656635&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22284906%26dopt%3DAbstract Authors: Symonds EL, Fenech M Abstract ABSTRACT: The APCmin/+ mouse is commonly used in cancer research and is just one of many genetically altered models that is currently being developed. With high numbers of breeding programs, it is important to have a simple method that can be used to genotype the mice non-invasively. Here we report a reproducible method for genotyping mice with DNA extracted from fecal samples. Comparison of fecal results with those obtained from intestinal tissue DNA and clinical outcome (presence/absence of tumors) showed this technique to have 100% accuracy. This non-invasive method of genotyping may be applied to other transgenic mouse models. PMID: 22284906 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5656635 Mon, 30 Jan 2012 05:00:00 +0100 5656635 http://www.medworm.com/index.php?rid=5377509&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22035318%26dopt%3DAbstract Authors: Mancini C, Messana E, Turco E, Brussino A, Brusco A Abstract ABSTRACT: In the preparation of transgenic murine ES cells it is important to verify the construct has a single insertion, because an ectopic neomycin phosphortransferase positive selection cassette (NEO) may cause a position effect. During a recent work, where a knockin SCA28 mouse was prepared, we developed two assays based on Real-Time PCR using both SYBR Green and specific minor groove binder (MGB) probes to evaluate the copies of NEO using the comparative delta-delta Ct method versus the Rpp30 reference gene. We compared the results from Southern blot, routinely used to quantify NEO copies, with the two Real-Time PCR assays. Twenty-two clones containing the single NEO copy showed values of 0.98 +/- 0.24 (mea... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5377509 Fri, 28 Oct 2011 04:00:00 +0100 5377509 http://www.medworm.com/index.php?rid=5343386&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D22011343%26dopt%3DAbstract Authors: Al-Khazraji BK, Medeiros PJ, Novielli NM, Jackson DN Abstract ABSTRACT: A cell-counting algorithm, developed in Matlab, was created to efficiently count migrated fluorescently-stained cells on membranes from migration assays. At each concentration of cells used (10,000, and 100,000 cells), images were acquired at 2.5x, 5x, and 10x objective magnifications. Automated cell counts strongly correlated to manual counts (r^2 = 0.99, P<0.0001 for a total of 47 images), with no difference in the measurements between methods under all conditions. We conclude that our automated method is accurate, more efficient, and void of variability and potential observer bias normally associated with manual counting. PMID: 22011343 [PubMed - as supplied by publisher] (Source: Biological ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5343386 Wed, 19 Oct 2011 04:00:00 +0100 5343386 http://www.medworm.com/index.php?rid=5324604&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21982381%26dopt%3DAbstract We describe here a method to rapidly convert any desirable DNA fragment, as small as 100 bp, into long tandem DNA arrays up to 140 kb in size that are inserted into a microbe vector. This method includes rolling-circle phi29 amplification (RCA) of the sequence in vitro and assembly of the RCA products in vivo by homologous recombination in the yeast Saccharomyces cerevisiae. The method was successfully used for a functional analysis of centromeric and pericentromeric repeats and construction of new vehicles for gene delivery to mammalian cells. The method may have general application in elucidating the role of tandem repeats in chromosome organization and dynamics. Each cycle of the protocol takes ~ two weeks to complete. PMID: 21982381 [PubMed - as supplied by publisher] (Source: Biol... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5324604 Fri, 07 Oct 2011 04:00:00 +0100 5324604 http://www.medworm.com/index.php?rid=5294420&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21970368%26dopt%3DAbstract Authors: Arvin F, Doraisamy S, Safar Khorasani E Abstract ABSTRACT: Auscultation is an approach for diagnosing many cardiovascular problems. Automatic analysis of heartbeat sounds and extraction of its audio features can assist physicians towards diagnosing diseases. Textual transcription allows recording a continuous heart sound stream using a text format which can be stored in very small memory in comparison with other audio formats. In addition, a text-based data allows applying indexing and searching techniques to access to the critical events. Hence, the transcribed heartbeat sounds provides useful information to monitor the behavior of a patient for the long duration of time. This paper proposes a frequency shifting method in order to improve the performance of the transcript... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5294420 Tue, 04 Oct 2011 04:00:00 +0100 5294420 http://www.medworm.com/index.php?rid=5139907&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21843345%26dopt%3DAbstract Authors: Liu Y, Merchant Z, Hsiao HC, Gonzalez KL, Matthews KS, Bondos SE Abstract ABSTRACT: Although yeast two-hybrid experiments are commonly used to identify protein interactions, the frequent occurrence of false negatives and false positives hampers data interpretation. Using both yeast one-hybrid and two-hybrid experiments, we have identified potential sources of these problems: the media preparation protocol and the source of the yeast nitrogen base may not only impact signal range but also effect whether a result appears positive or negative. While altering media preparation may optimize signal differences for individual experiments, media preparation must be reported in detail to replicate studies and accurately compare results from different experiments. PMID: 21843345... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5139907 Sun, 14 Aug 2011 23:00:00 +0100 5139907 http://www.medworm.com/index.php?rid=5092847&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21777412%26dopt%3DAbstract Authors: Mseddi F, Jarboui MA, Sellami A, Sellami H, Ayadi A ABSTRACT: DNA isolation from C. neoformans is difficult due to a thick and resistant capsule. We have optimized a new and rapid DNA isolation method for Cryptococcus using a short urea treatment followed by a rapid method using a chelex resin suspension. This procedure considerably simplifies previously reported methods. PMID: 21777412 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=5092847 Wed, 20 Jul 2011 23:00:00 +0100 5092847 http://www.medworm.com/index.php?rid=4949846&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21663682%26dopt%3DAbstract Conclusions Digital image analysis can be used for screening agents for potential impact on reproductive toxicity or carcinogenesis as well as for mechanistic studies, particularly for cumulative effects on mammary epithelial mass as well as translational studies of mechanisms that explain the relationship between epithelial mass and cancer risk. PMID: 21663682 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=4949846 Thu, 09 Jun 2011 23:00:00 +0100 4949846 http://www.medworm.com/index.php?rid=4575296&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21369532%26dopt%3DAbstract Authors: Chen G, Kronenberger P, Teugels E, De Grève J Real-time quantitative RT-PCR (RT-qPCR) is a "gold" standard for measuring steady state mRNA levels in RNA interference assays. The knockdown of the epidermal growth factor receptor (EGFR) gene with eight individual EGFR small interfering RNAs (siRNAs) was estimated by RT-qPCR using three different RT-qPCR primer sets. PMID: 21369532 [PubMed - in process] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=4575296 Sat, 12 Mar 2011 07:00:03 +0100 4575296 http://www.medworm.com/index.php?rid=4575295&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21369533%26dopt%3DAbstract Authors: Li S PMID: 21369533 [PubMed - in process] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=4575295 Sat, 12 Mar 2011 07:00:03 +0100 4575295 http://www.medworm.com/index.php?rid=4575294&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D21369534%26dopt%3DAbstract We describe a simple and reproducible method to measure absolute telomere length (aTL) using quantitative real-time polymerase chain reaction (qPCR). This method is based on the Cawthon method for relative measurement of telomere length (TL) but modified by introducing an oligomer standard to measure aTL. The method describes the oligomer standards, the generation of the standard curve and the calculations required to calculate aTL from the qPCR data. The necessary controls and performance characteristics of the assay are described in detail and compared relative to other methods for measuring TL. Typical results for this assay for a variety of human tissue samples are provided as well as a troubleshooting schedule. This method allows high throughput measurement of aTL using small amounts ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=4575294 Sat, 12 Mar 2011 07:00:03 +0100 4575294 http://www.medworm.com/index.php?rid=3057258&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19957062%26dopt%3DAbstract Authors: Carter JD, Dula SB, Corbin KL, Wu R, Nunemaker CS Pancreatic islets of Langerhans secrete hormones that are vital to the regulation of blood glucose and are, therefore, a key focus of diabetes research. Purifying viable and functional islets from the pancreas for study is an intricate process. This review highlights the key elements involved with mouse and rat islet isolation, including choices of collagenase, the collagenase digestion process, purification of islets using a density gradient, and islet culture conditions. In addition, this paper reviews commonly used techniques for assessing islet viability and function, including visual assessment, fluorescent markers of cell death, glucose-stimulated insulin secretion, and intracellular calcium measurements. A detailed proto... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=3057258 Thu, 03 Dec 2009 00:00:00 +0100 3057258 http://www.medworm.com/index.php?rid=3057257&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19957063%26dopt%3DAbstract Authors: Maioli E, Torricelli C, Fortino V, Carlucci F, Tommassini V, Pacini A Rottlerin is a natural product isolated from Mallotus philippinensis. This polyphenolic compound, originally described as a selective inhibitor of PKCdelta, can inhibit many other PKC-unrelated kinases and has a number of biological actions, including mitochondrial uncoupling effects. We recently found that Rottlerin inhibits the transcription factor nuclear factor kappaB in different cell types, causing downregulation of cyclin D1 and growth arrest. The present study was carried out to clarify the surprising lack of effect of Rottlerin on MCF-7 cell viability, assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) test. We found that Rottlerin causes overestimation of the MTT te... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=3057257 Thu, 03 Dec 2009 00:00:00 +0100 3057257 http://www.medworm.com/index.php?rid=3057256&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19957064%26dopt%3DAbstract Authors: Gothelf A, Hojman P, Gehl J Analgesic and anesthetic drugs may have an impact on the results achieved from animal experiments. In the study presented here, we try to enlighten whether anesthesia with fentanyl/fluniasone and midazolam (Hypnorm and Dormicum) has an influence on measurements of hemoglobin in mice. In a cross-over study, we have compared hemoglobin levels in two groups of mice: anesthetized versus non-anesthetized and found significant decrease in hemoglobin levels in the anesthetized group (p < 0.05) unrelated to which group received the anesthesia. The mean hemoglobin levels after intraperitoneal administration of Hypnorm and Dormicum was 8.7 mmol/L compared to mean hemoglobin 9.9 mmol/L before anesthesia (p < 0.001), and the decrease lasted for more than ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=3057256 Thu, 03 Dec 2009 00:00:00 +0100 3057256 http://www.medworm.com/index.php?rid=3057255&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19957065%26dopt%3DAbstract Authors: Zhang L, Ren F, Li J, Ma X, Wang P In order to determine the activity of O-linked GlcNAc transferase (OGT), a modified coupled enzyme method was proposed. This method was based on the measurement of uridine 5'-(trihydrogen diphosphate) (UDP), a product generated in transglycosylation reaction. In the assay, UDP was coupled to the conversion of phosphoenolpyruvate to pyruvate using pyruvate kinase. Using a commercial pyruvate assay kit, the pyruvate was converted to a red terminal product, which could be photometrically measured at 570 nm or fluorometrically measured at 587 nm (E (m) = 535 nm) on a microplate reader. Kinetic study of a truncated recombinant mOGT and quantitative analysis of OGT in two biological samples indicated that this method was practical and competitive f... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=3057255 Thu, 03 Dec 2009 00:00:00 +0100 3057255 http://www.medworm.com/index.php?rid=2804818&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19756912%26dopt%3DAbstract In this study, we report on the application of a real-time noninvasive system for dynamic monitoring of cellular plasticity. Analysis of the cell impedance profile recorded as cell index using a real-time cell analyzer revealed its significant increase after the treatment of prostate epithelial cells with the transforming growth factor-beta1. Changes in the cell index profile were paralleled with cytoskeleton rebuilding and induction of epithelial-mesenchymal transition and negatively correlated with cell proliferation. This novel application of such approach demonstrated a great potential of the impedance-based system for noninvasive and real-time monitoring of cellular fate. PMID: 19756912 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2804818 Fri, 11 Sep 2009 23:00:00 +0100 2804818 http://www.medworm.com/index.php?rid=2766860&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19728006%26dopt%3DAbstract Authors: Li S PMID: 19728006 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2766860 Mon, 31 Aug 2009 23:00:00 +0100 2766860 http://www.medworm.com/index.php?rid=2640099&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19629613%26dopt%3DAbstract Authors: Siegrist F, Singer T, Certa U MicroRNAs are positive and negative regulators of eukaryotic gene expression that modulate transcript abundance by specific binding to sequence motifs located prevalently in the 3' untranslated regions of target messenger RNAs (mRNA). Interferon-alpha-2a (IFNalpha) induces a large set of protein coding genes mediating antiproliferative and antiviral responses. Here we use a global microarray-based microRNA detection platform to identify genes that are induced by IFNalpha in hepatoma- or melanoma-derived human tumor cell lines. Despite the enormous differences in expression levels between these models, we were able to identify microRNAs that are upregulated by IFNalpha in both lines suggesting the possibility that interferon-regulated microRNAs are... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2640099 Wed, 22 Jul 2009 23:00:00 +0100 2640099 http://www.medworm.com/index.php?rid=2602818&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19597903%26dopt%3DAbstract In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effective approach for ectopic transgene expression and an alternative to generation of transgenic animals. One benefit to this approach is that it can be used independently or in conjunction with established transgenic or knockout animals for studying modulation of mesenchymal-epithelial interactions. To display the power of this approach, we explored ectopic expression of a Wnt ligand in the mouse intestinal mesenchyme and demonstrate its functional influence on the adjacent epithelium. Our findings highlight the efficient use of lentiviral-mediated transgene expression for modulating mesenchymal-epithelial interactions in vivo. PMID: 19597903 [PubMed - as supplied by publisher] (Source: Biological ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2602818 Mon, 13 Jul 2009 23:00:00 +0100 2602818 http://www.medworm.com/index.php?rid=2602817&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19597904%26dopt%3DAbstract Authors: Avazpour I, Saripan MI, Nordin AJ, Abdullah RS In the image segmentation process of positron emission tomography combined with computed tomography (PET/CT) imaging, previous works used information in CT only for segmenting the image without utilizing the information that can be provided by PET. This paper proposes to utilize the hot spot values in PET to guide the segmentation in CT, in automatic image segmentation using seeded region growing (SRG) technique. This automatic segmentation routine can be used as part of automatic diagnostic tools. In addition to the original initial seed selection using hot spot values in PET, this paper also introduces a new SRG growing criterion, the sliding windows. Fourteen images of patients having extrapulmonary tuberculosis have been exami... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2602817 Mon, 13 Jul 2009 23:00:00 +0100 2602817 http://www.medworm.com/index.php?rid=2548377&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19551458%26dopt%3DAbstract Authors: Pappas JJ, Toulouse A, Bradley WE The bisulfite genomic sequencing protocol is a widely used method for analyzing DNA methylation. It relies on the deamination of unmethylated cytosine residues to uracil; however, its high rates of DNA degradation and incomplete cytosine to uracil conversion often lead to failed experiments, uninformative results, and false positives. Here, we report the addition of a single-step multiple restriction enzyme digestion (MRED) designed to differentially digest polymerase chain reaction products amplified from unconverted DNA while leaving those of converted DNA intact. We show that for our model system, RARB2 P2 promoter, use of MRED increased informative sequencings ninefold, and MRED did not alter the clonal representation in one fully methylat... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548377 Tue, 23 Jun 2009 23:00:00 +0100 2548377 http://www.medworm.com/index.php?rid=2548381&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19517206%26dopt%3DAbstract Authors: Sadeghian F, Seman Z, Ramli AR, Abdul Kahar BH, Saripan MI Evaluation of blood smear is a commonly clinical test these days. Most of the time, the hematologists are interested on white blood cells (WBCs) only. Digital image processing techniques can help them in their analysis and diagnosis. For example, disease like acute leukemia is detected based on the amount and condition of the WBC. The main objective of this paper is to segment the WBC to its two dominant elements: nucleus and cytoplasm. The segmentation is conducted using a proposed segmentation framework that consists of an integration of several digital image processing algorithms. Twenty microscopic blood images were tested, and the proposed framework managed to obtain 92% accuracy for nucleus segmentation and 78% f... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548381 Wed, 10 Jun 2009 23:00:00 +0100 2548381 http://www.medworm.com/index.php?rid=2548379&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19517207%26dopt%3DAbstract Authors: Tian X, Syvitski RT, Liu T, Livingstone N, Jakeman DL, Li YH Many species of streptococci secrete and use a competence-stimulating peptide (CSP) to initiate quorum sensing for induction of genetic competence, bacteriocin production, and other activities. These signaling molecules are small, unmodified peptides that induce powerful strain-specific activity at nano-molar concentrations. This feature has provided an excellent opportunity to explore their structure-function relationships. However, CSP variants have also been identified in many species, and each specifically activates its cognate receptor. How such minor changes dramatically affect the specificity of these peptides remains unclear. Structure-activity analysis of these peptides may provide clues for understanding th... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548379 Wed, 10 Jun 2009 23:00:00 +0100 2548379 http://www.medworm.com/index.php?rid=2548400&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495910%26dopt%3DAbstract Authors: Erickson HP An important part of characterizing any protein molecule is to determine its size and shape. Sedimentation and gel filtration are hydrodynamic techniques that can be used for this medium resolution structural analysis. This review collects a number of simple calculations that are useful for thinking about protein structure at the nanometer level. Readers are reminded that the Perrin equation is generally not a valid approach to determine the shape of proteins. Instead, a simple guideline is presented, based on the measured sedimentation coefficient and a calculated maximum S, to estimate if a protein is globular or elongated. It is recalled that a gel filtration column fractionates proteins on the basis of their Stokes radius, not molecular weight. The molecular we... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548400 Thu, 14 May 2009 23:00:00 +0100 2548400 http://www.medworm.com/index.php?rid=2548398&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495911%26dopt%3DAbstract Authors: Wai-Hoe L, Wing-Seng L, Ismail Z, Lay-Harn G Kidney stone disease is a common health problem in industrialised nations. We developed a SDS-PAGE-based method to quantify Tamm Horsfall glycoprotein (THP) for screening of kidney stone disease. Urinary proteins were extracted by using ammonium sulphate precipitation at 0.27 g salt/mL urine. The resulted pellet was dissolved in TSE buffer. Ten microliters of the urinary proteins extract was loaded and separated on 10% SDS-PAGE under reducing condition. THP migrated as single band in SDS-PAGE. The assay reproducibility and repeatability were 4.8% CV and 2.6% CV, respectively. A total of 117 healthy subjects and 58 stone patients were tested using this assay, and a distinct cut-off (P < 0.05) at 5.6 mug/mL THP concentration was us... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548398 Thu, 14 May 2009 23:00:00 +0100 2548398 http://www.medworm.com/index.php?rid=2548394&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495913%26dopt%3DAbstract Authors: Hojman P, Eriksen J, Gehl J DNA electrotransfer to muscle tissue yields long-term, high levels of gene expression; showing great promise for future gene therapy. We want to characterize the novel far-red fluorescent protein Katushka as a marker for gene expression using time domain fluorescence in vivo imaging. Highly efficient transgenic expression was observed after DNA electrotransfer with 100-fold increase in fluorescent intensity. The fluorescent signal peaked 1 week after transfection and returned to background level within 4 weeks. Katushka expression was not as stable as GFP expression, which was detectable for 8 weeks. Depth and 3D analysis proved that the expression was located in the target muscle. In vivo bio-imaging using the novel Katushka fluorescent protein ena... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548394 Thu, 09 Apr 2009 23:00:00 +0100 2548394 http://www.medworm.com/index.php?rid=2548395&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495912%26dopt%3DAbstract Authors: Hasan MA, Reaz MB, Ibrahimy MI, Hussain MS, Uddin J Fetal electrocardiogram (FECG) signal contains potentially precise information that could assist clinicians in making more appropriate and timely decisions during labor. The ultimate reason for the interest in FECG signal analysis is in clinical diagnosis and biomedical applications. The extraction and detection of the FECG signal from composite abdominal signals with powerful and advance methodologies are becoming very important requirements in fetal monitoring. The purpose of this review paper is to illustrate the various methodologies and developed algorithms on FECG signal detection and analysis to provide efficient and effective ways of understanding the FECG signal and its nature for fetal monitoring. A comparative stud... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548395 Fri, 27 Mar 2009 00:00:00 +0100 2548395 http://www.medworm.com/index.php?rid=2548391&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495914%26dopt%3DAbstract Authors: Commins J, Toft C, Fares MA Comparative genomics has become a real tantalizing challenge in the postgenomic era. This fact has been mostly magnified by the plethora of new genomes becoming available in a daily bases. The overwhelming list of new genomes to compare has pushed the field of bioinformatics and computational biology forward toward the design and development of methods capable of identifying patterns in a sea of swamping data noise. Despite many advances made in such endeavor, the ever-lasting annoying exceptions to the general patterns remain to pose difficulties in generalizing methods for comparative genomics. In this review, we discuss the different tools devised to undertake the challenge of comparative genomics and some of the exceptions that compromise the ge... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548391 Wed, 11 Mar 2009 00:00:00 +0100 2548391 http://www.medworm.com/index.php?rid=2548387&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495916%26dopt%3DAbstract Authors: Rodríguez-Mallon A, Cárdenas Y, Lugo JM, Oliva A, Morales A, Estrada MP Quantization of gene expression requires that an accurate measurement of a specific transcript is made. In this paper, a quantitative reverse transcription-polymerase chain reaction (RT-PCR) by competition for tilapia growth hormone receptor type I is designed and validated. This experimental procedure was used to determine the abundance of growth hormone receptor type I transcript in different tilapia tissues. The results obtained with this developed competitive RT-PCR were similar to real-time PCR results reported recently. This protocol provides a reliable alternative, but less expensive than real-time PCR to quantify specific genes. PMID: 19495916 [PubMed - as supplied by publisher] (Sour... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548387 Tue, 10 Mar 2009 00:00:00 +0100 2548387 http://www.medworm.com/index.php?rid=2548385&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495917%26dopt%3DAbstract Authors: Heng BC, Cowan CM, Basu S The dissociation of adherent mesenchymal stem cell (MSC) monolayers with trypsin and enzyme-free dissociation buffer was compared. A significantly lower proportion of viable cells were obtained with enzyme-free dissociation buffers compared to trypsin. Subsequently, the dissociated cells were re-seeded on new cell culture dishes and were subjected to the MTT assay 24 h later. The proportion of viable cells that reattached was significantly lower for cells obtained by dissociation with enzyme-free dissociation buffer compared to trypsin. Frozen-thawed MSC displayed a similar trend, yielding consistently higher cell viability and reattachment rates when dissociated with trypsin compared to enzyme-free dissociation buffer. It was also demonstrated that e... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548385 Tue, 10 Mar 2009 00:00:00 +0100 2548385 http://www.medworm.com/index.php?rid=2548389&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495915%26dopt%3DAbstract We describe an optimized method to speed up the preparation of suitable testicular cell suspensions for cytometric analysis of different spermatogenic stages from rodents. The procedure takes only 15 min including testis dissection, tissue cutting, and processing through the Medimachine System (Becton Dickinson). This method could be a substitute for the more tedious and time-consuming cell preparation techniques currently in use. PMID: 19495915 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548389 Fri, 06 Mar 2009 00:00:00 +0100 2548389 http://www.medworm.com/index.php?rid=2548384&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19495918%26dopt%3DAbstract Authors: Motyl K, McCabe LR As many as 50% of adults with type I (T1) diabetes exhibit bone loss and are at increased risk for fractures. Therapeutic development to prevent bone loss and/or restore lost bone in T1 diabetic patients requires knowledge of the molecular mechanisms accounting for the bone pathology. Because cell culture models alone cannot fully address the systemic/metabolic complexity of T1 diabetes, animal models are critical. A variety of models exist including spontaneous and pharmacologically induced T1 diabetic rodents. In this paper, we discuss the streptozotocin (STZ)-induced T1 diabetic mouse model and examine dose-dependent effects on disease severity and bone. Five daily injections of either 40 or 60 mg/kg STZ induce bone pathologies similar to spontaneously di... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2548384 Fri, 06 Mar 2009 00:00:00 +0100 2548384 http://www.medworm.com/index.php?rid=2007846&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D19048127%26dopt%3DAbstract In this report, we detail the different approaches used for the isolation, the identification of a RNA binding protein and the validation of its role. PMID: 19048127 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=2007846 Wed, 03 Dec 2008 18:01:25 +0100 2007846 http://www.medworm.com/index.php?rid=1591637&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385804%26dopt%3DAbstract Authors: Morales H, Robert J Activation of lymphocytes in mammals is often quantified by measuring the amount of proliferation during the expansion phase of an immune response. Bromodeoxyuridine (BrdU) incorporation and carboxyfluorescein diacetate succinimidyl ester (CFSE) dilution assays are some of the techniques widely used in mammalian studies of pathogen-induced proliferation and provide a convenient way of quantifying the cellular response. We have extended the use of these proliferation assays to the amphibian Xenopus laevis. We have developed this species as a valuable comparative model to study immunity against a well-known amphibian pathogen, Frog Virus 3 (FV3). Fluorescence activated cell sorting was used to assess the level of BrdU incorporation of lymphocytes in vivo and ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591637 Mon, 07 Jul 2008 20:42:01 +0100 1591637 http://www.medworm.com/index.php?rid=1591636&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385805%26dopt%3DAbstract Authors: Moeenrezakhanlou A, Nandan D, Reiner NE Calcitriol (1alpha, 25-dihydroxyvitamin D(3)) induces the expression of CD14 in mononuclear phagocytes. The mechanisms accounting for this have been unclear since the promoter of CD14 does not contain a canonical vitamin D response element (VDRE). Calcitriol has been shown to regulate the activity of the transcription factor Sp-1 and our analysis of the proximal promoter of CD14 indicated the presence of four Sp-1-like binding sequences. To identify which of these sites might be involved in the response to calcitriol, we used a system incorporating an electrophoretic mobility shift assay (EMSA) coupled to Western blot analysis (WEMSA). Using WEMSA, we found that only one of the Sp-1-like binding sequences, located at position -91 to -79 ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591636 Mon, 07 Jul 2008 20:42:01 +0100 1591636 http://www.medworm.com/index.php?rid=1591635&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385806%26dopt%3DAbstract Authors: Keating DH, Shulla A, Klein AH, Wolfe AJ Acetyl phosphate (acetyl-P) serves critical roles in coenzyme A recycling and ATP synthesis. It is the intermediate of the Pta-AckA pathway that inter-converts acetyl-coenzyme A and acetate. Acetyl-P also can act as a global signal by donating its phosphoryl group to specific two-component response regulators. This ability derives from its capacity to store energy in the form of a high-energy phosphate bond. This bond, while critical to its function, also destabilizes acetyl-P in cell extracts. This lability has greatly complicated biochemical analysis, leading in part to widely varying acetyl-P measurements. We therefore developed an optimized protocol based on two-dimensional thin layer chromatography that includes metabolic labeling ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591635 Mon, 07 Jul 2008 20:42:01 +0100 1591635 http://www.medworm.com/index.php?rid=1591634&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385807%26dopt%3DAbstract Authors: Robenek H, Severs NJ Freeze-fracture electron microscopy is a technique for examining the ultrastructure of rapidly frozen biological samples by transmission electron microscopy. Of a range of approaches to freeze-fracture cytochemistry that have been developed and tried, the most successful is the technique termed freeze-fracture replica immunogold labeling (FRIL). In this technique, samples are frozen, fractured and replicated with platinum-carbon as in standard freeze fracture, and then carefully treated with sodium dodecylsulphate to remove all the biological material except a fine layer of molecules attached to the replica itself. Immunogold labeling of these molecules permits their distribution to be seen superimposed upon high resolution planar views of membrane structu... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591634 Mon, 07 Jul 2008 20:42:01 +0100 1591634 http://www.medworm.com/index.php?rid=1591633&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385808%26dopt%3DAbstract In this report, we describe a simple method for the quantification of JNK activation by Western blot analysis or directly in tissue culture plates. PMID: 18385808 [PubMed - in process] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591633 Mon, 07 Jul 2008 20:42:01 +0100 1591633 http://www.medworm.com/index.php?rid=1591644&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17364022%26dopt%3DAbstract Authors: Murphy M, Greferath U, Wilson YM We have developed a system to visualize functionally activated neurons and their projections in the brain. This system utilizes a transgenic mouse, fos-tau-lacZ (FTL), which expresses the marker gene, lacZ, in neurons and their processes after activation by many different stimuli. This system allows the imaging of activation from the level of the entire brain surface, through to individual neurons and their projections. The use of this system involves detection of neuronal activation by histochemical or immunohistochemical detection of beta-galactosidase (betagal), the product of the lacZ gene. Furthermore, the underlying brain state of the FTL mice determines the basal levels of expression of betagal. Here we describe in detail our protocols f... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591644 Mon, 01 Jan 2007 05:00:00 +0100 1591644 http://www.medworm.com/index.php?rid=1591643&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18213360%26dopt%3DAbstract Authors: Wu H, Dinh A, Mo YY Suppression of gene expression by small interfering RNA (siRNA) has proved to be a gene-specific and cost effective alternative to other gene suppression technologies. Short hairpin RNAs (shRNAs) generated from the vector-based expression are believed to be processed into functional siRNAs in vivo, leading to gene silencing. Since an shRNA library carries a large pool of potential siRNAs, such a library makes it possible to knock down gene expression at the genome wide scale. Although much of research has been focused on generating shRNA libraries from either individually made gene specific sequences or cDNA libraries, there is no report on constructing randomized shRNA libraries, which could provide a good alternative to these existing libraries. We have d... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591643 Mon, 01 Jan 2007 05:00:00 +0100 1591643 http://www.medworm.com/index.php?rid=1591642&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18213361%26dopt%3DAbstract Authors: Deng Q, Luo W, Donnenberg MS We developed a rapid mutagenesis method based on a modification of the QuikChange(R) system (Stratagene) to systemically replace endogenous gene sequences with a unique similar size sequence tag. The modifications are as follows: 1: the length of the anchoring homologous sequences of both mutagenesis primers were increased to 16 - 22 bp to achieve melting temperatures greater than 80 degrees C. 2: the final concentrations of both primers were increased to 5-10 ng/mul and the final concentration of template to 1-2 ng/mul. 3: the annealing temperature was adjusted when necessary from 52 degrees C to 58 degrees C. We generated 25 sequential mutants in the cloned espD gene (1.2 kb), which encodes an essential component of the type III secretion translo... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591642 Mon, 01 Jan 2007 05:00:00 +0100 1591642 http://www.medworm.com/index.php?rid=1591641&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18213362%26dopt%3DAbstract Authors: Koo SH, Ong TC, Chong KT, Lee CG, Chew FT, Lee EJ We have developed and validated a consolidated bead-based genotyping platform, the Bioplex suspension array for simultaneous detection of multiple single nucleotide polymorphisms (SNPs) of the ATP-binding cassette transporters. Genetic polymorphisms have been known to influence therapeutic response and risk of disease pathologies. Genetic screening for therapeutic and diagnostic applications thus holds great promise in clinical management. The allele-specific primer extension (ASPE) reaction was used to assay 22 multiplexed SNPs for eight subjects. Comparison of the microsphere-based ASPE assay results to sequencing results showed complete concordance in genotype assignments. The Bioplex suspension array thus proves to be a rel... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591641 Mon, 01 Jan 2007 05:00:00 +0100 1591641 http://www.medworm.com/index.php?rid=1591640&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18213363%26dopt%3DAbstract Authors: Salinas I, Meseguer J, Esteban MA Teleost gut associated lymphoid tissue (GALT) consists of leucocyte populations located both intraepithelially and in the lamina propria with no structural organization. The present study aims to assess different protocols for the isolation of GALT cells from an important fish species in the Mediterranean aquaculture, the gilthead seabream. Mechanical, chemical and enzymatic treatments were assayed. Nylon wool columns and continuous density gradients were used for further separation of cell subpopulations. Light microscopy and flow cytometry showed that the highest density band (HD) consisted of a homogeneous lymphocytic population, whereas the intermediate density band (ID) corresponded to epithelial and secretory cells and some lymphocytes. ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591640 Mon, 01 Jan 2007 05:00:00 +0100 1591640 http://www.medworm.com/index.php?rid=1591639&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18335004%26dopt%3DAbstract Authors: Patel N, Castillo M, Rameshwar P In adults, hematopoiesis occurs in bone marrow (BM) through a complex process with differentiation of hematopoietic stem cells (HSCs) to immune and blood cells. Human HSCs and their progenitors express CD34. Methods on hematopoietic regulation are presented to show the effects of the chemokine, stromal-derived growth factor (SDF)-1I and the neuropeptide, substance P (SP). SDF-1I production in BM stroma causes interactions with HSCs, thereby retaining the HSCs in regions close to the endosteum, at low oxygen. Small changes in SDF-1I levels stimulate HSC functions through direct and indirect mechanisms. The indirect method occurs by SP production, which stimulates CD34+ cells, supported by ligand-binding studies, long-term culture-initiating cell... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591639 Mon, 01 Jan 2007 05:00:00 +0100 1591639 http://www.medworm.com/index.php?rid=1591638&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385803%26dopt%3DAbstract In this study, we present a new method which allows detection of protein oxidation and total protein on the same gel to improve matching in image analysis. Furthermore, the digested protein spots are compatible with standard MALDI mass spectrometry protein identification. The methodology highlighted here may be useful in facilitating the development of biomarkers, assessing potential therapeutic targets and elucidating new mechanisms of redox signalling in redox-related conditions. PMID: 18385803 [PubMed - in process] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591638 Mon, 01 Jan 2007 05:00:00 +0100 1591638 http://www.medworm.com/index.php?rid=1591632&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18385809%26dopt%3DAbstract Authors: Antony P, Hoek K, Sarmah B, Khan WN B cell subpopulations in the spleen have been extensively characterized phenotypically; however, biochemical properties of these cell populations following B cell antigen receptor engagement have not been fully determined due to technical difficulties and limiting cell numbers. We therefore employed mini-scale protocols to assess lipid signaling, particularly that of diacylglycerol and inositol trisphosphate, with as few as 0.5x10(6) purified early (T1) and late (T2) transitional B cells. Additionally, utilizing flow cytometric techniques, we determined levels of phosphatidylinositol bisphosphate and calcium mobilization in T1 and T2 cells, as well as mature follicular and marginal zone B cells using less than 1x10(6 )primary B cells. Thus, ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591632 Mon, 01 Jan 2007 05:00:00 +0100 1591632 http://www.medworm.com/index.php?rid=1591631&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D18464937%26dopt%3DAbstract Authors: Fernandez SL, Russell DW, Hurlin PJ Understanding mechanisms of gene regulation has broad therapeutic implications for human disease. Here we describe a novel method for generating human cell lines that serve as reporters of transcriptional activity. This method exploits the ability of recombinant adeno-associated virus to mediate the insertion of exogenous DNA sequences into specific genomic loci through homologous recombination. To overcome the severe size limitation of the rAAV for carrying exogenous DNA, an enhanced green fluorescent protein (EGFP)-Luciferase fusion gene was used as both a selectable marker and gene expression reporter. EGFP was used for selection of correctly targeted alleles by taking advantage of known regulatory conditions that activate transcription o... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591631 Mon, 01 Jan 2007 05:00:00 +0100 1591631 http://www.medworm.com/index.php?rid=1591657&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16446781%26dopt%3DAbstract We describe in this paper a simple protocol which uses real-time PCR to compare mRNA levels of a gene of interest between different experimental conditions. Comparative real-time PCR can be a relatively low-cost method and does not require sequence-specific fluorescent reporters. Moreover, several genes from a set of experiments can be assessed in a single run. Thus, in addition to providing a comparative profile for the expression of a gene of interest, this method can also provide information regarding the relative abundance of different mRNA species. PMID: 16446781 [PubMed] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591657 Sun, 01 Jan 2006 05:00:00 +0100 1591657 http://www.medworm.com/index.php?rid=1591656&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16799694%26dopt%3DAbstract Authors: Raez MB, Hussain MS, Mohd-Yasin F Electromyography (EMG) signals can be used for clinical/biomedical applications, Evolvable Hardware Chip (EHW) development, and modern human computer interaction. EMG signals acquired from muscles require advanced methods for detection, decomposition, processing, and classification. The purpose of this paper is to illustrate the various methodologies and algorithms for EMG signal analysis to provide efficient and effective ways of understanding the signal and its nature. We further point up some of the hardware implementations using EMG focusing on applications related to prosthetic hand control, grasp recognition, and human computer interaction. A comparison study is also given to show performance of various EMG signal analysis methods. This ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591656 Sun, 01 Jan 2006 05:00:00 +0100 1591656 http://www.medworm.com/index.php?rid=1591655&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16799695%26dopt%3DAbstract Authors: Yamada HY, Gorbsky GJ Microtubule inhibitors such as Vinblastine and Paclitaxel are chemotherapy agents that activate the mitotic spindle checkpoint, arresting cells in mitosis and leading to cell death. The pathways that connect mitotic arrest to cell death are not well characterized. We developed a mammalian cell-based cDNA cloning method to isolate proteins and protein fragments whose expression inhibits colony formation in the presence of microtubule inhibitors. Understanding how these proteins impact cellular responses to microtubule drugs will lead to better understanding of the biochemical pathways connecting mitotic arrest and cell death in mammalian cells and may provide novel targets that can enhance microtubule inhibitor-mediated chemotherapy. PMID: 16799695 [Pu... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591655 Sun, 01 Jan 2006 05:00:00 +0100 1591655 http://www.medworm.com/index.php?rid=1591654&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16799696%26dopt%3DAbstract In this report, we demonstrate the selective in vivo association of NF-kappaB, p300 and CREB with the human Igamma1 promoter located in the intronic region upstream of the Cgamma1 exons in the immunoglobulin heavy chain locus. These methods have the ability to extend ChIP analysis to promoters with a high degree of homology. PMID: 16799696 [PubMed] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591654 Sun, 01 Jan 2006 05:00:00 +0100 1591654 http://www.medworm.com/index.php?rid=1591653&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16900260%26dopt%3DAbstract Authors: Austin BA, James CM, Härle P, Carr DJ The application of naked DNA containing type I interferon (IFN) transgenes is a promising potential therapeutic approach for controlling chronic viral infections. Herein, we detail the application of this approach that has been extensively used to restrain ocular HSV-1 infection, for antagonizing vaginal HSV-2 infection. We show that application of IFN-alpha1, -alpha5, and -beta transgenes to vaginal mouse lumen 24 hours prior to HSV-2 infection reduces HSV-2 mediated mortality by 2.5 to 3-fold. However, other type I IFN transgenes (IFN- alpha4, -alpha5, -alpha6, and -alpha9) are non effectual against HSV-2. We further show that the efficacy of IFN-alpha1 transgene treatment is independent of CD4+ T lymphocytes. However, in mice deple... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591653 Sun, 01 Jan 2006 05:00:00 +0100 1591653 http://www.medworm.com/index.php?rid=1591652&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16909160%26dopt%3DAbstract Authors: Xia J, Kim SH, Macmillan S, Truant R Live cell fluorescence microscopy using fluorescent protein tags derived from jellyfish and coral species has been a successful tool to image proteins and dynamics in many species. Multi-colored aequorea fluorescent protein (AFP) derivatives allow investigators to observe multiple proteins simultaneously, but overlapping spectral properties sometimes require the use of sophisticated and expensive microscopes. Here, we show that the aequorea coerulescens fluorescent protein derivative, PS-CFP2 has excellent practical properties as a blue fluorophore that are distinct from green or red fluorescent proteins and can be imaged with standard filter sets on a widefield microscope. We also find that by widefield illumination in live cells, that PS-... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591652 Sun, 01 Jan 2006 05:00:00 +0100 1591652 http://www.medworm.com/index.php?rid=1591651&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17033696%26dopt%3DAbstract Authors: Emmerson JR, Gally DL, Roe AJ In this work we describe protocols for the generation of gene deletions and gene replacements using a temperature sensitive plasmid in Escherichia coli O157:H7. This technology requires flanking DNA to be cloned into a temperature sensitive vector but the resulting clone allows great flexibility for further modification of the target sequence. It is therefore highly suited to the study of genes in which several rounds of changes are envisaged. A number of examples are used to illustrate the flexibility of the system which has been used to create novel gene replacements including fusions for protein localisation work and reporters for transcriptional analyses. In this paper we describe protocols which can be used with a high degree of success when ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591651 Sun, 01 Jan 2006 05:00:00 +0100 1591651 http://www.medworm.com/index.php?rid=1591650&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17033697%26dopt%3DAbstract Authors: Ray S, Das SK Chromatin immunoprecipitation (ChIP) technique allows detection of proteins that bind to chromatin. While this technique has been applied extensively in cell-based studies, its tissue-based application remains poorly explored. We are specifically interested in examining estrogen-dependent transcriptional mechanism in respect of recruitment of estrogen receptor-alpha (ERalpha), a ligand-activated transcription factor, to uterine gene promoters in mice. Recent gene-array studies, utilizing ERalpha knock-out vs. wild-type mice, have revealed that estrogen regulates numerous uterine genes temporally and most importantly via ERalpha during the phase-II response, including three well characterized genes viz., lactoferrin (Ltf), progesterone receptor (Pgr) and cyclinD1 ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591650 Sun, 01 Jan 2006 05:00:00 +0100 1591650 http://www.medworm.com/index.php?rid=1591649&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17033698%26dopt%3DAbstract Authors: Badtke MP, Cao F, Tavis JE Protein-protein interactions are required for many viral and cellular functions and are potential targets for novel therapies. Here we detail a series of genetic and biochemical techniques used in combination to find an essential molecular contact point on the duck hepatitis B virus polymerase. These techniques include differential immunoprecipitation, mutagenesis and peptide competition. The strength of these techniques is their ability to identify contact points on intact proteins or protein complexes employing functional assays. This approach can be used to aid identification of putative binding sites on proteins and protein complexes which are resistant to characterization by other methods. PMID: 17033698 [PubMed] (Source: Biological Procedur... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591649 Sun, 01 Jan 2006 05:00:00 +0100 1591649 http://www.medworm.com/index.php?rid=1591648&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17033699%26dopt%3DAbstract Authors: Gallup JM, Ackermann MR The purpose of this manuscript is to discuss fluorogenic real-time quantitative polymerase chain reaction (qPCR) inhibition and to introduce/define a novel Microsoft Excel-based file system which provides a way to detect and avoid inhibition, and enables investigators to consistently design dynamically-sound, truly LOG-linear qPCR reactions very quickly. The qPCR problems this invention solves are universal to all qPCR reactions, and it performs all necessary qPCR set-up calculations in about 52 seconds (using a pentium 4 processor) for up to seven qPCR targets and seventy-two samples at a time - calculations that commonly take capable investigators days to finish. We have named this custom Excel-based file system "FocusField2-6GallupqPCRSet-upTool-001"... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591648 Sun, 01 Jan 2006 05:00:00 +0100 1591648 http://www.medworm.com/index.php?rid=1591647&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17066123%26dopt%3DAbstract Authors: Lin M, Zhao W, Wu R How the power required for bird flight varies as a function of forward speed can be used to predict the flight style and behavioral strategy of a bird for feeding and migration. A U-shaped curve was observed between the power and flight velocity in many birds, which is consistent to the theoretical prediction by aerodynamic models. In this article, we present a general genetic model for fine mapping of quantitative trait loci (QTL) responsible for power curves in a sample of birds drawn from a natural population. This model is developed within the maximum likelihood context, implemented with the EM algorithm for estimating the population genetic parameters of QTL and the simplex algorithm for estimating the QTL genotype-specific parameters of power curves. ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591647 Sun, 01 Jan 2006 05:00:00 +0100 1591647 http://www.medworm.com/index.php?rid=1591646&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17242735%26dopt%3DAbstract Authors: Morey JS, Ryan JC, Van Dolah FM Quantitative real-time PCR (qPCR) is a commonly used validation tool for confirming gene expression results obtained from microarray analysis; however, microarray and qPCR data often result in disagreement. The current study assesses factors contributing to the correlation between these methods in five separate experiments employing two-color 60-mer oligonucleotide microarrays and qPCR using SYBR green. Overall, significant correlation was observed between microarray and qPCR results (rho=0.708, p<0.0001, n=277) using these platforms. The contribution of factors including up- vs. down-regulation, spot intensity, rho-value, fold-change, cycle threshold (C(t)), array averaging, tissue type, and tissue preparation was assessed. Filtering of micr... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591646 Sun, 01 Jan 2006 05:00:00 +0100 1591646 http://www.medworm.com/index.php?rid=1591645&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D17242736%26dopt%3DAbstract Authors: Harrison JJ, Ceri H, Yerly J, Stremick CA, Hu Y, Martinuzzi R, Turner RJ Microbes frequently live within multicellular, solid surface-attached assemblages termed biofilms. These microbial communities have architectural features that contribute to population heterogeneity and consequently to emergent cell functions. Therefore, three-dimensional (3D) features of biofilm structure are important for understanding the physiology and ecology of these microbial systems. This paper details several protocols for scanning electron microscopy and confocal laser scanning microscopy (CLSM) of biofilms grown on polystyrene pegs in the Calgary Biofilm Device (CBD). Furthermore, a procedure is described for image processing of CLSM data stacks using amira, a virtual reality tool, to create su... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591645 Sun, 01 Jan 2006 05:00:00 +0100 1591645 http://www.medworm.com/index.php?rid=1591671&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15678169%26dopt%3DAbstract Authors: Smart N, Scambler PJ, Riley PR RNA Interference has rapidly emerged as an efficient procedure for knocking down gene expression in model systems. However, cross-reactivity, whereby multiple genes may be simultaneously targeted by a single short interfering RNA (siRNA), can potentially jeopardize correct interpretation of gene function. As such, it is essential to test the specificity of a siRNA prior to a full phenotypic analysis. To this end, we have adapted a reporter-based assay harnessing the sensitivity of luciferase activity to provide a quantitative readout of relative RNAi efficacy and specificity. We have tested different siRNAs directed against Thymosin beta4 (Tbeta4); determined their effectiveness at silencing Tbeta4 and have both excluded off-target silencing of t... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591671 Sat, 01 Jan 2005 05:00:00 +0100 1591671 http://www.medworm.com/index.php?rid=1591670&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15678170%26dopt%3DAbstract This study describes a reliable and reproducible method for isolating and expanding malignant cells from surgical breast tissues. The method uses co-cultures with BM stroma to select for the cancer cells while the healthy cells undergo rapid cell death. Studies are described to show the cloning efficiencies and sensitivity of the method using surgical samples of varying sizes, different stages of BC, and samples from needle biopsies. PMID: 15678170 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591670 Sat, 01 Jan 2005 05:00:00 +0100 1591670 http://www.medworm.com/index.php?rid=1591669&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15682222%26dopt%3DAbstract Authors: Lefrancois S, Canuel M, Zeng J, Morales CR To assess the role of sortilin in the sorting and trafficking of sphingolipid activator proteins (SAPs) the function of sortilin was abolished by a dominant-negative mutant and by the use of RNAi. Mutant sortilin lacking the carboxyl-terminal region that contains the sorting signal abolished the trafficking of SAPs to the lysosomes. Both sortilin and SAPs were retained in the Golgi apparatus. The use of chemically synthesized siRNA effectively blocked the trafficking of SAPs to the lysosomes as well. Additionally, we created a stable COS-7 cell line transfected with the pSilencer 3.1 H1 neo vector containing a selected siRNA template oligonucleotide (small hairpin interference RNA) where the levels of sortilin were greatly suppressed.... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591669 Sat, 01 Jan 2005 05:00:00 +0100 1591669 http://www.medworm.com/index.php?rid=1591668&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136221%26dopt%3DAbstract In this study we have observed that A72(dog), CRFK(cat), NB324K(human), MCF7(human), WI38(human), and C636(mosquito) cells were effectively cryopreserved at -80 degrees C while attached to the substratum of 25 cm2 tissue culture flasks. This was accomplished using a serum free crypreservative recently developed by Corsini and co-workers. The technique allows for significant savings of time and money in laboratories that rapidly process numerous cell lines. PMID: 16136221 [PubMed] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591668 Sat, 01 Jan 2005 05:00:00 +0100 1591668 http://www.medworm.com/index.php?rid=1591667&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136222%26dopt%3DAbstract We describe an improved, efficient and reliable method for the vapour-phase silanization of multi-barreled, ion-selective microelectrodes of which the silanized barrel(s) are to be filled with neutral liquid ion-exchanger (LIX). The technique employs a metal manifold to exclusively and simultaneously deliver dimethyldichlorosilane to only the ion-selective barrels of several multi-barreled microelectrodes. Compared to previously published methods the technique requires fewer procedural steps, less handling of individual microelectrodes, improved reproducibility of silanization of the selected microelectrode barrels and employs standard borosilicate tubing rather than the less-conventional theta-type glass. The electrodes remain stable for up to 3 weeks after the silanization procedure. The... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591667 Sat, 01 Jan 2005 05:00:00 +0100 1591667 http://www.medworm.com/index.php?rid=1591666&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136223%26dopt%3DAbstract Authors: Watanabe M, Fujioka-Kaneko Y, Kobayashi H, Kiniwa M, Kuwano M, Basaki Y Angiogenesis is a complex process involving an ECM and vascular endothelial cells (EC), and is regulated by various angiogenic factors including VEGF. The ability to form a capillary/tube-like network is a specialized function of EC. Therefore, in vitro angiogenesis was assessed by a capillary/tube-like network formation assay. There are three angiogenic parameters: capillary length, number of capillaries, and relative capillary area per field. We evaluated capillary length per field in the assay. VEGF promoted capillary/tube-like network formation of EC in a type I collagen gel matrix in vitro. Moreover, we demonstrated the involvement of ILK in a VEGF signaling pathway mediating capillary/tube-like netwo... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591666 Sat, 01 Jan 2005 05:00:00 +0100 1591666 http://www.medworm.com/index.php?rid=1591665&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136224%26dopt%3DAbstract Authors: Akiyama T, Miyazaki T, Bouillet P, Nakamura K, Strasser A, Tanaka S Mature osteoclasts, multinucleated giant cells responsible for bone resorption, are terminally differentiated cells with a short life span. Recently, we have demonstrated that osteoclast apoptosis is regulated by ERK activity and Bcl-2 family member Bim. In this paper, we summarize the methods we used to study osteoclast apoptosis in vitro and in vivo. Using adenovirus and retrovirus vectors, we were able to introduce foreign genes into osteoclasts and examine their effects on osteoclast survival in vitro. In addition, we established the modified methods for in situ hybridization and BrdU labeling of bone sections from mice to study osteoclast survival in vivo. The detailed methods described here could be usef... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591665 Sat, 01 Jan 2005 05:00:00 +0100 1591665 http://www.medworm.com/index.php?rid=1591664&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136225%26dopt%3DAbstract Authors: Ricke RM, Bielinsky AK The Histone Association Assay provides an easy approach for detecting proteins that bind chromatin in vivo. This technique is based on a chromatin immunoprecipitation protocol using histone H3-specific antibodies to precipitate bulk chromatin from crosslinked whole cell extracts. Proteins that co-precipitate with chromatin are subsequently detected by conventional SDS-PAGE and Western blot analysis. Unlike techniques that separate chromatin and non-chromatin interacting proteins by centrifugation, this method can be used to delineate whether a protein is chromatin associated regardless of its innate solubility. Moreover, the relative amount of protein bound to DNA can be ascertained under quantitative conditions. Therefore, this technique may be utilized... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591664 Sat, 01 Jan 2005 05:00:00 +0100 1591664 http://www.medworm.com/index.php?rid=1591663&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136226%26dopt%3DAbstract We describe a new approach for reliably isolating one-step real-time quantitative RT-PCR-quality RNA from laser captured cells retrieved from frozen sections previously subjected to immunofluorescent immunohistochemistry (IF-IHC) and subsequently subjected to fluorogenic one-step real-time RT-PCR analysis without the need for costly, time-consuming linear amplification. One cell's worth of RNA can now be interrogated with confidence. This approach represents an amalgam of technologies already offered commercially by Applied Biosystems, Arcturus and Invitrogen. It is the primary focus of this communication to expose the details and execution of an important new LCM RNA isolation technique, but also provide a detailed account of the IF-IHC procedure preceding RNA isolation, and provide infor... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591663 Sat, 01 Jan 2005 05:00:00 +0100 1591663 http://www.medworm.com/index.php?rid=1591662&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136227%26dopt%3DAbstract Authors: Chandra PK, Wikel SK We have developed a simple and effective method (Lig-PCR) for monitoring ligation reactions using PCR and primers that are common to many cloning vectors. Ligation mixtures can directly be used as templates and the results can be analyzed by conventional gel electrophoresis. The PCR products are representative of the recombinant molecules created during ligation and the corresponding transformants. Orientation of inserts can also be determined using an internal primer. The usefulness of this method has been demonstrated using ligation mixtures of two cDNA's derived from the salivary glands of Aedes aegypti mosquitoes. The method described here is sensitive and easy to perform compared to currently available methods. PMID: 16136227 [PubMed] (Source: Bio... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591662 Sat, 01 Jan 2005 05:00:00 +0100 1591662 http://www.medworm.com/index.php?rid=1591661&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16136228%26dopt%3DAbstract Authors: Jud C, Schmutz I, Hampp G, Oster H, Albrecht U Most behavioral experiments within circadian research are based on the analysis of locomotor activity. This paper introduces scientists to chronobiology by explaining the basic terminology used within the field. Furthermore, it aims to assist in designing, carrying out, and evaluating wheel-running experiments with rodents, particularly mice. Since light is an easily applicable stimulus that provokes strong effects on clock phase, the paper focuses on the application of different lighting conditions. PMID: 16136228 [PubMed] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591661 Sat, 01 Jan 2005 05:00:00 +0100 1591661 http://www.medworm.com/index.php?rid=1591660&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16145578%26dopt%3DAbstract Authors: Encarnación S, Hernández M, Martínez-Batallar G, Contreras S, Vargas Mdel C, Mora J We propose two-dimensional gel electrophoresis (2-DE) and mass spectrometry to define the protein components of regulons and stimulons in bacteria, including those organisms where genome sequencing is still in progress. The basic 2-DE protocol allows high resolution and reproducibility and enables the direct comparison of hundreds or even thousands of proteins simultaneously. To identify proteins that comprise stimulons and regulons, peptide mass fingerprint (PMF) with matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF-MS) analysis is the first option and, if results from this tool are insufficient, complementary data obtained with electrospra... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591660 Sat, 01 Jan 2005 05:00:00 +0100 1591660 http://www.medworm.com/index.php?rid=1591659&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16281079%26dopt%3DAbstract In this study we develop an experimental design to sensitize pancreatic islet cells by high glucose to streptozotocin (STZ) and proinflammatory cytokines [interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma]-induced apoptosis. This method is appropriate for subsequent quantification of apoptotic islet cells stained with Tdt-mediated dUTP Nick-End Labeling (TUNEL) and protein expression assays by Western Blotting (WB). PMID: 16281079 [PubMed] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591659 Sat, 01 Jan 2005 05:00:00 +0100 1591659 http://www.medworm.com/index.php?rid=1591658&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D16299584%26dopt%3DAbstract Authors: Wozniak MA, Keely PJ Several pathological and disease conditions can alter the mechanical properties of the extracellular matrix (ECM). Conversely, some diseases may arise from changes in the density or rigidity of the ECM. This necessitates the use and development of in vitro models to understand how both biophysical and biochemical signals regulate complex cellular behaviors. T47D breast epithelial cells will differentiate into duct-like tubules when cultured in a floating three-dimensional (3D) collagen gel, but not a 3D collagen gel that is left attached to the culture dish. This paper details several protocols we have developed for analyzing breast cell biology in 3D matrices, including culturing cells in 3D collagen gels, immunostaining cellular structures, and performin... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591658 Sat, 01 Jan 2005 05:00:00 +0100 1591658 http://www.medworm.com/index.php?rid=1591702&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14737221%26dopt%3DAbstract We describe the use of atomic force microscopy (AFM) in studies of cell adhesion and cell compliance. Our studies use the interaction between leukocyte function associated antigen-1 (LFA-1)/intercellular adhesion molecule-1 (ICAM-1) as a model system. The forces required to unbind a single LFA-1/ICAM-1 bond were measured at different loading rates. This data was used to determine the dynamic strength of the LFA-1/ICAM-1 complex and characterize the activation potential that this complex overcomes during its breakage. Force measurements acquired at the multiple- bond level provided insight about the mechanism of cell adhesion. In addition, the AFM was used as a microindenter to determine the mechanical properties of cells. The applications of these methods are described using data from a pr... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591702 Thu, 01 Jan 2004 05:00:00 +0100 1591702 http://www.medworm.com/index.php?rid=1591701&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103395%26dopt%3DAbstract Authors: Zaragoza O, Casadevall A Experimental modulation of capsule size is an important technique for the study of the virulence of the encapsulated pathogen Cryptococcus neoformans. In this paper, we summarize the techniques available for experimental modulation of capsule size in this yeast and describe improved methods to induce capsule size changes. The response of the yeast to the various stimuli is highly dependent on the cryptococcal strain. A high CO(2) atmosphere and a low iron concentration have been used classically to increase capsule size. Unfortunately, these stimuli are not reliable for inducing capsular enlargement in all strains. Recently we have identified new and simpler conditions for inducing capsule enlargement that consistently elicited this effect. Specificall... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591701 Thu, 01 Jan 2004 05:00:00 +0100 1591701 http://www.medworm.com/index.php?rid=1591700&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103396%26dopt%3DAbstract Authors: Firulli BA, Virshup DM, Firulli AB Post-translational modifications such as phosphorylation play a vital role in the regulation of protein function. In our study of the basic Helix-loop-Helix (bHLH) transcription factor HAND1, it was suspected that HAND1 was being phosphorylated during trophoblast giant cell differentiation and that coexpression of a constitutively active kinase with HAND1 resulted in changes in the proteins dimerization profile. In order to accurately document HAND1 phosphorylation and identify the resides being modified, we employed metabolic cell labeling with (32)P of tissue culture cells coexpressing a Flag-epitope tagged HAND1 along with a number of active kinases and phosphatase subunits. We generated phosphopeptide maps of the phosphorylated HAND1 usin... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591700 Thu, 01 Jan 2004 05:00:00 +0100 1591700 http://www.medworm.com/index.php?rid=1591699&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103397%26dopt%3DAbstract Authors: Ohi M, Li Y, Cheng Y, Walz T Vitrification is the state-of-the-art specimen preparation technique for molecular electron microscopy (EM) and therefore negative staining may appear to be an outdated approach. In this paper we illustrate the specific advantages of negative staining, ensuring that this technique will remain an important tool for the study of biological macromolecules. Due to the higher image contrast, much smaller molecules can be visualized by negative staining. Also, while molecules prepared by vitrification usually adopt random orientations in the amorphous ice layer, negative staining tends to induce preferred orientations of the molecules on the carbon support film. Combining negative staining with image classification techniques makes it possible to work wi... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591699 Thu, 01 Jan 2004 05:00:00 +0100 1591699 http://www.medworm.com/index.php?rid=1591698&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103398%26dopt%3DAbstract Authors: Antoun A, Pavlov MY, Tenson T, Ehrenberg M Light scattering and standard stopped-flow techniques were used to monitor rapid association of ribosomal subunits during initiation of eubacterial protein synthesis. The effects of the initiation factors IF1, IF2, IF3 and buffer conditions on subunit association were studied along with the role of GTP in this process. The part of light scattering theory that is essential for kinetic measurements is high-lighted in the main text and a more general treatment of Rayleigh scattering from macromolecules is given in an appendix. PMID: 15103398 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591698 Thu, 01 Jan 2004 05:00:00 +0100 1591698 http://www.medworm.com/index.php?rid=1591697&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103399%26dopt%3DAbstract Authors: Stark HJ, Szabowski A, Fusenig NE, Maas-Szabowski N To assess the role of genes required for skin organogenesis, tissue regeneration and homeostasis, we have established in vitro skin equivalents composed of primary cells or cell lines, respectively. In these organotypic cocultures keratinocytes generate a normal epidermis irrespective of the species and tissue origin of fibroblasts. The combination of cells derived from mouse and human tissues facilitates the identification of the origin of compounds involved in epidermal tissue reconstitution and thus the precise analysis of growth regulatory mechanisms. PMID: 15103399 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591697 Thu, 01 Jan 2004 05:00:00 +0100 1591697 http://www.medworm.com/index.php?rid=1591696&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15103400%26dopt%3DAbstract Authors: Corsini J, Hacker C, Bare C Herein we have explored two practical aspects of cryopreserving cultured mammalian cells during routine laboratory maintenance. First, we have examined the possibility of using a serum-free, hence more affordable, cryopreservative. Using five mammalian lines (Crandell Feline Kidney, MCF7, A72, WI 38 and NB324K), we found that the serum-free alternative preserves nearly as efficiently as the serum-containing preservatives. Second, we compared cryostorage of those cells in suspended versus a pellet form using both aforementioned cryopreservatives. Under our conditions, cells were in general recovered equally well in a suspended versus a pellet form. PMID: 15103400 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591696 Thu, 01 Jan 2004 05:00:00 +0100 1591696 http://www.medworm.com/index.php?rid=1591695&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15181476%26dopt%3DAbstract Authors: Munir S, Singh S, Kaur K, Kapur V High throughput detection of differential expression of genes is an efficient means of identifying genes and pathways that may play a role in biological systems under certain experimental conditions. There exist a variety of approaches that could be used to identify groups of genes that change in expression in response to a particular stimulus or environment. We here describe the application of suppression subtractive hybridization (SSH) coupled with cDNA microarray analysis for isolation and identification of chicken transcripts that change in expression on infection of host cells with a paramyxovirus. SSH was used for initial isolation of differentially expressed transcripts, a large-scale validation of which was accomplished by microarray a... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591695 Thu, 01 Jan 2004 05:00:00 +0100 1591695 http://www.medworm.com/index.php?rid=1591694&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15181477%26dopt%3DAbstract Authors: Manna PR, Huhtaniemi IT, Stocco DM The steroidogenic acute regulatory (StAR) protein, a novel mitochondrial protein, is involved in the regulation of steroid hormone biosynthesis through its mediation of the intramitochondrial transport of the steroid substrate, cholesterol, to the cytochrome P450 cholesterol side chain cleavage (P450scc) enzyme. The expression of StAR protein is regulated by cAMP-dependent signaling in steroidogenic cells. During the course of our studies in mouse Leydig cells, we employ several methods for studying the regulation of StAR protein expression by human chorionic gonadotropin (hCG). A sensitive quantitative reverse transcription and polymerase chain reaction (RT-PCR) was utilized for determining StAR mRNA expression. Stimulation of mLTC-1 mouse L... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591694 Thu, 01 Jan 2004 05:00:00 +0100 1591694 http://www.medworm.com/index.php?rid=1591693&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15188014%26dopt%3DAbstract Authors: Centonze VE, Firulli BA, Firulli AB Post-translational modifications such as phosphorylation play a vital role in the regulation of protein function. In our study of the basic Helix-loop-Helix (bHLH) transcription factor HAND1, we show that HAND1 is phosphorylated during the trophoblast giant cell differentiation on residues residing in Helix I of the bHLH domain. Our hypothesis is that these modifications result in changes in HAND1 dimerization affinities with other bHLH factors. To test this idea, we employed FRET to measure the protein-protein interactions of HAND1 and HAND1 point mutants in HEK293 cells using YFP and CFP fusion proteins and laser scanning confocal microscopy. PMID: 15188014 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591693 Thu, 01 Jan 2004 05:00:00 +0100 1591693 http://www.medworm.com/index.php?rid=1591692&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15188015%26dopt%3DAbstract Authors: Li J, Zhang WB, McManus DP Three cDNAs, termed EpC1, TPxEg and EgG5, were isolated by immunoscreening from an Echinococcus granulosus cDNA library. The recombinant phages exhibited strong reactivity with sera from humans with confirmed cystic echinococcosis (CE) and with sera from mice infected with E. granulosus oncospheres. The cDNAs were subcloned into a pET vector, expressed as fusion proteins tagged with GST and affinity purified against the GST tag. Of the three recombinant proteins, EpC1 achieved the highest performance for serodiagnosis of CE in Western blot analysis using a panel of clinically defined human sera to initially address the sensitivity and specificity of the molecules. The protein yielded an overall sensitivity of 92.2% and specificity of 95.6%, levels un... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591692 Thu, 01 Jan 2004 05:00:00 +0100 1591692 http://www.medworm.com/index.php?rid=1591691&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15192754%26dopt%3DAbstract Authors: Davies AG, McIntire SL Caenorhabditis elegans is an attractive model system for determining the targets of neuroactive compounds. Genetic screens in C. elegans provide a relatively unbiased approach to the identification of genes that are essential for behavioral effects of drugs and neuroactive compounds such as alcohol. Much work in vertebrate systems has identified multiple potential targets of ethanol but which, if any, of those candidates are responsible for the behavioral effects of alcohol is uncertain. Here we provide detailed methodology for a genetic screen for mutants of C. elegans that are resistant to the depressive effects of ethanol on locomotion and for the subsequent behavioral analysis of those mutants. The methods we describe should also be applicable for us... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591691 Thu, 01 Jan 2004 05:00:00 +0100 1591691 http://www.medworm.com/index.php?rid=1591690&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15192755%26dopt%3DAbstract Authors: Andersen SS In synaptic vesicles, the estimated concentration of the excitatory amino acid glutamate is 100-150 mM. It was recently discovered that VGLUT1, previously characterized as an inorganic phosphate transporter (BNPI) with 9-11 predicted transmembrane spanning domains, is capable of transporting glutamate. The expression and His-tag based purification of recombinant VGLUT1 from PC12 cells and High Five insect cells is described. Significantly better virus and protein expression was obtained using High Five rather than Sf9 insect cells. PC12 cell expressed VGLUT1 is functional but not the Baculovirus expressed protein. The lack of functionality of the Baculovirus expressed VGLUT1 is discussed. The data indicate that VGLUT1 readily oligomerizes/dimerizes. The data are di... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591690 Thu, 01 Jan 2004 05:00:00 +0100 1591690 http://www.medworm.com/index.php?rid=1591689&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15243647%26dopt%3DAbstract Authors: Iqbal Z, Siddiqui RT, Qureshi JA Imatinib (Gleevec) is the effective therapy for BCR-ABL positive CML patients. Point mutations have been detected in ATP-binding domain of ABL gene which disturbs the binding of Gleevec to this target leading to resistance. Detection of mutations is helpful in clinical management of imatinib resistance. We established a very sensitive (ASO) PCR to detect mutations in an imatinib-resistant CML patient. Mutations C944T and T1052C were detected which cause complete partial imatinib resistance, respectively. This is the first report of multiple point mutations conferring primary imatinib resistance in same patient at the same time. Understanding the biological reasons of primary imatinib resistance is one of the emerging issues of pharmacogenomics ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591689 Thu, 01 Jan 2004 05:00:00 +0100 1591689 http://www.medworm.com/index.php?rid=1591688&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15243648%26dopt%3DAbstract Authors: Pomati F, Burns BP, Neilan BA Here we describe an experimental design aimed to investigate changes in total cellular levels of Na(+) and K(+) ions in cultures of freshwater filamentous cyanobacteria. Ion concentrations were measured in whole cells by flame photometry. Cellular Na(+) levels increased exponentially with rising alkalinity, with K(+) levels being maximal for optimal growth pH (~8). At standardized pH conditions, the increase in cellular Na(+), as induced by NaCl at 10 mM, was coupled by the two sodium channel-modulating agents lidocaine hydrochloride at 1 microM and veratridine at 100 microM. Both the channel-blockers amiloride (1 mM) and saxitoxin (1 microM), decreased cell-bound Na(+) and K(+) levels. Results presented demonstrate the robustness of well-defined ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591688 Thu, 01 Jan 2004 05:00:00 +0100 1591688 http://www.medworm.com/index.php?rid=1591687&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15243649%26dopt%3DAbstract Authors: Chaves-Pozo E, Mulero V, Meseguer J, García Ayala A The gilthead seabream is a protandrous seasonal breeding teleost that is an excellent model for studying the testicular regression process which occurs in both seasonal testicular involution and sex reversion. Little is known about the cell types and the molecular mechanisms involved in such processes, mainly because of the lack of appropriate methods for testis dissociation, and testicular cell isolation, culture and functional characterization. We have previously reported that gilthead seabream acidophilic granulocytes infiltrate the testis at post-spawning stage, settle close to the spermatogonia and accumulate intracellular interleukin-1beta. In this paper, we report several flow cytometry based assays which allow to... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591687 Thu, 01 Jan 2004 05:00:00 +0100 1591687 http://www.medworm.com/index.php?rid=1591686&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15243650%26dopt%3DAbstract Authors: Kienberger F, Zhu R, Moser R, Rankl C, Blaas D, Hinterdorfer P Dynamic force microscopy (DFM) allows imaging of the structure and the assessment of the function of biological specimens in their physiological environment. In DFM, the cantilever is oscillated at a given frequency and touches the sample only at the end of its downward movement. Accordingly, the problem of lateral forces displacing or even destroying bio-molecules is virtually inexistent as the contact time and friction forces are reduced. Here, we describe the use of DFM in studies of human rhinovirus serotype 2 (HRV2) weakly adhering to mica surfaces. The capsid of HRV2 was reproducibly imaged without any displacement of the virus. Release of the genomic RNA from the virions was initiated by exposure to low pH b... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591686 Thu, 01 Jan 2004 05:00:00 +0100 1591686 http://www.medworm.com/index.php?rid=1591685&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15272365%26dopt%3DAbstract Authors: Gerhart J, Baytion M, Perlman J, Neely C, Hearon B, Nilsen T, Getts R, Kadushin J, George-Weinstein M In situ hybridization with 3DNA trade mark dendrimers is a novel tool for detecting low levels of mRNA in tissue sections and whole embryos. Fluorescently labeled dendrimers were used to identify cells that express mRNA for the skeletal muscle transcription factor MyoD in the early chick embryo. A small population of MyoD mRNA positive cells was found in the epiblast prior to the initiation of gastrulation, two days earlier than previously detected using enzymatic or radiolabeled probes for mRNA. When isolated from the epiblast and placed in culture, the MyoD mRNA positive cells were able to differentiate into skeletal muscle cells. These results demonstrate that DNA dendrimer... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591685 Thu, 01 Jan 2004 05:00:00 +0100 1591685 http://www.medworm.com/index.php?rid=1591684&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15282629%26dopt%3DAbstract Authors: Vermeulen M, Stunnenberg HG Post-translational modifications of core histones play an important role in regulating fundamental biological processes such as DNA repair, transcription and replication. In this paper, we describe a novel assay that allows sequential targeting of distinct histone modifying enzymes to immobilized nucleosomal templates using recombinant chimeric targeting molecules. The assay can be used to study the histone substrate specificity of chromatin modifying enzymes as well as whether and how certain enzymes affect each other's histone modifying activities. As such the assay can help to understand how a certain histone code is established and interpreted. PMID: 15282629 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591684 Thu, 01 Jan 2004 05:00:00 +0100 1591684 http://www.medworm.com/index.php?rid=1591683&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15328539%26dopt%3DAbstract Authors: Pinhero R, Liaw P, Yankulov K We have established a uniform procedure for the expression and purification of the cyclin-dependent kinases CDK7/CycH/MAT1, CDK8/CycC and CDK9/CycT1. We attach a His(6)-tag to one of the subunits of each complex and then co-express it together with the other subunits in Spodoptera frugiperda insect cells. The CDK complexes are subsequently purified by Ni(2+)-NTA and Mono S chromatography. This approach generates large amounts of active recombinant kinases that are devoid of contaminating kinase activities. Importantly, the properties of these recombinant kinases are similar to their natural counterparts (Pinhero et al. 2004, Eur J Biochem 271:1004-14). Our protocol provides a novel systematic approach for the purification of these three (and possi... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591683 Thu, 01 Jan 2004 05:00:00 +0100 1591683 http://www.medworm.com/index.php?rid=1591682&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15346173%26dopt%3DAbstract Authors: Dunne JL, Goobic AP, Acton ST, Ley K Leukocyte endothelial cell interaction is a fundamentally important process in many disease states. Current methods to analyze such interactions include the parallel-plate flow chamber and intravital microscopy. Here, we present an improvement of the traditional intravital microscopy that allows leukocyte-endothelial cell interaction to be studied from the time the leukocyte makes its initial contact with the endothelium until it adheres to or detaches from the endothelium. The leukocyte is tracked throughout the venular tree with the aid of a motorized stage and the rolling and adhesive behavior is measured off-line. Because this method can involve human error, methods to automate the tracking procedure have been developed. This novel trac... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591682 Thu, 01 Jan 2004 05:00:00 +0100 1591682 http://www.medworm.com/index.php?rid=1591681&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15361931%26dopt%3DAbstract We describe how such analyses can be performed using existing phylogenetic tools, and discuss how new phylogenetic analysis tools we have recently developed can be used to provide more detailed and sensitive analyses, including study of the evolution of mutation and substitution processes. As an example we consider the mitochondrial genome, for which two types of transition deaminations (C-->T and A-->G) are strongly affected by single-strandedness during replication, resulting in a strand asymmetric mutation process. Since time spent single-stranded varies along the mitochondrial genome, their differential mutational response results in very different substitution patterns in different regions of the genome. PMID: 15361931 [PubMed - as supplied by publisher] (Source: Biological ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591681 Thu, 01 Jan 2004 05:00:00 +0100 1591681 http://www.medworm.com/index.php?rid=1591680&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15448721%26dopt%3DAbstract We reported that DNA methylation and chromatin structure contributes to lymphoid-specific ITGAL (CD11a) and PRF1 (perforin) expression. We used bisulfite sequencing to compare methylation patterns in the ITGAL promoter and 5' flanking region of T cells and fibroblasts, and in the PRF1 promoter and upstream enhancer of CD4+ and CD8+ T cells with fibroblasts. The effects of methylation on promoter function were tested using regional methylation of reporter constructs, and confirmed by DNA methyltransferase inhibition. The relationship between DNA methylation and chromatin structure was analyzed by DNaseI hypersensitivity. Herein we described the methods and results in greater detail. PMID: 15448721 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591680 Thu, 01 Jan 2004 05:00:00 +0100 1591680 http://www.medworm.com/index.php?rid=1591679&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15472720%26dopt%3DAbstract Authors: Li J, Sha B We have identified a peptide substrate for molecular chaperone Hsp40 Ydj1 by utilizing the combination of phage display library screening and isothemol titration calirimetry (ITC). The initial peptide substrate screening for Hsp40 Ydj1 has been carried out by utilizing a 7-mer phage display library. The peptide sequences from the bio-panning were synthesized and object to the direct affinity measurement for Hsp40 Ydj1 by isothemol titration calirimetry studies. The peptide which has the measurable affinity with Ydj1 shows enriched hydrophobic residues in the middle of the substrate fragment. The peptide substrate specificity for molecular chaperone Hsp40 has been analyzed. PMID: 15472720 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591679 Thu, 01 Jan 2004 05:00:00 +0100 1591679 http://www.medworm.com/index.php?rid=1591678&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15472721%26dopt%3DAbstract Authors: Shirley RL, Richards MR, Culbertson MR A method was developed to assess the functional significance of a sequence motif in yeast Upf3p, a protein required for nonsense-mediated mRNA decay (NMD). The motif lies at the edge of the Upf3p-Upf2p interaction domain, but at the same time resembles the canonical leucine-rich nuclear export sequence (NES) found in proteins that bind Crm1p exportin. To test the function of the putative NES, site-directed mutations that cause substitutions of conserved NES-A residues were first selected to identify hypermorphic alleles. Next, a portable Crm1p-binding NES from HIV-1 Rev protein that functions in yeast was fused en masse to the C-terminus of variant Upf3 proteins using loxP sites recognized by bacterial cre-recombinase. Finally, variant Up... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591678 Thu, 01 Jan 2004 05:00:00 +0100 1591678 http://www.medworm.com/index.php?rid=1591677&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15472722%26dopt%3DAbstract Authors: Pick N, Cameron S, Arad D, Av-Gay Y The worldwide rapid increase in bacterial resistance to numerous antibiotics requires on-going development of new drugs to enter the market. As the development of new antibiotics is lengthy and costly, early monitoring of compound's toxicity is essential in the development of novel agents. Our interest is in a rapid, simple, high throughput screening method to assess cytotoxicity induced by potential agents. Some intracellular pathogens, such as Mycobacterium tuberculosis primary site of infection is human alveolar macrophages. Thus, evaluation of candidate drugs for macrophage toxicity is crucial. Protocols for high throughput drug toxicity screening of macrophages using flow cytometry are lacking in the literature. For this application we ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591677 Thu, 01 Jan 2004 05:00:00 +0100 1591677 http://www.medworm.com/index.php?rid=1591676&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15514698%26dopt%3DAbstract Authors: Rommelaere H, Waterschoot D, Neirynck K, Vandekerckhove J, Ampe C Recombinant production and biochemical analysis of actin mutants has been hampered by the fact that actin has an absolute requirement for the eukaryotic chaperone CCT to reach its native state. We therefore have developed a method to rapidly screen the folding capacity and functionality of actin variants, by combining in vitro expression of labelled actin with analysis on native gels, band shift assays or copolymerization tests. Additionally, we monitor, using immuno-fluorescence, incorporation of actin variants in cytoskeletal structures in transfected cells. We illustrate the method by two examples. In one we show that tagged versions of actin do not always behave native-like and in the other we study some of ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591676 Thu, 01 Jan 2004 05:00:00 +0100 1591676 http://www.medworm.com/index.php?rid=1591675&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15514699%26dopt%3DAbstract Authors: Shinohara ET, Hallahan DE, Lu B Elucidating the mechanism of over and under expression of proteins is critical in developing a better understanding of cancer. Multiple techniques are used to examine differential expression of proteins in cells and assess changes in protein expression in response to therapies such as radiation. Reduced expression can be caused by protein inactivation, mRNA instability, or reduced transcription. The following protocol was used to determine the mechanism for the reduced expression of an antiapoptotic factor, survivin, in normal tissues in response to radiation and the defect in cancer cells that prevents this reduction. We also examined ways to overcome survivin over expression in cancer cells in order to sensitize them to radiation. We will focu... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591675 Thu, 01 Jan 2004 05:00:00 +0100 1591675 http://www.medworm.com/index.php?rid=1591674&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15562298%26dopt%3DAbstract Authors: Pan B, Shi K, Sundaralingam M Guanine-rich RNA oligonucleotides display many novel structural motifs in recent crystal structures. Here we describe the procedures of the chemical synthesis and the purification of such RNA molecules that are suitable for X-ray crystallographic studies. Modifications of the previous purification methods allow us to obtain better yields in shorter time. We also provide 24 screening conditions that are very effective in crystallization of the guanine-rich RNA oligonucleotides. Optimal crystallization conditions are usually achieved by adjustment of the concentration of the metal ions and pH of the buffer. Crystals obtained by this method usually diffract to high resolution. PMID: 15562298 [PubMed - as supplied by publisher] (Source: Biological... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591674 Thu, 01 Jan 2004 05:00:00 +0100 1591674 http://www.medworm.com/index.php?rid=1591673&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15605107%26dopt%3DAbstract Authors: Garimella R, Sipe JB, Anderson HC A simple and non-radioactive technique based on O-cresolpthalein complexone assay was developed to study in vitro non-radioactive calcium ((40)Ca) deposition by isolated matrix vesicles. Using this technique, the effect of various phosphoester substrates including ATP, AMP and beta-GP on in vitro MV-calcification was studied. O-cresolpthalein complexone assay with non-radioactive calcium demonstrated that AMP or beta-GP were more effective in promoting calcium deposition by isolated MVs than ATP. The application of this non-radioactive technique, which is highly sensitive and simple, would offer a useful alternative approach to the routinely used radiometric biomineralization assay which employs radioactive (45)Ca. PMID: 15605107 [PubMed -... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591673 Thu, 01 Jan 2004 05:00:00 +0100 1591673 http://www.medworm.com/index.php?rid=1591672&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D15630481%26dopt%3DAbstract We describe here a method for examining QS signaling systems in mammalian cells that uses engineered LuxR-type proteins from the opportunistic pathogen, Pseudomonas aeruginosa, which can function as AHL-dependent transcription factors. The engineered proteins respond to their cognate ligands and display sequence specific DNA binding properties. This system has several potential biotechnological and biological applications. It may be used to characterize any LuxR-type protein, screen animal and plant cell extracts or exudates for compounds that mimic or interfere with AHL signaling or to screen different cell types for AHL inactivating activities. PMID: 15630481 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591672 Thu, 01 Jan 2004 05:00:00 +0100 1591672 http://www.medworm.com/index.php?rid=1591717&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569610%26dopt%3DAbstract Authors: Uribe S, Sampedro JG In proteins, some processes require conformational changes involving structural domain diffusion. Among these processes are protein folding, unfolding and enzyme catalysis. During catalysis some enzymes undergo large conformational changes as they progress through the catalytic cycle. According to Kramers theory, solvent viscosity results in friction against proteins in solution, and this should result in decreased motion, inhibiting catalysis in motile enzymes. Solution viscosity was increased by adding increasing concentrations of glycerol, sucrose and trehalose, resulting in a decrease in the reaction rate of the H(+)-ATPase from the plasma membrane of Kluyveromyces lactis. A direct correlation was found between viscosity (eta) and the inhibition of the... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591717 Wed, 01 Jan 2003 05:00:00 +0100 1591717 http://www.medworm.com/index.php?rid=1591716&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569611%26dopt%3DAbstract Authors: Räthel TR, Leikert J , Vollmar AM, Dirsch VM Here we describe in more depth the previously published application of the fluorescent probe 4,5-diaminofluorescein (DAF-2) in order to reliably measure low levels of nitric oxide (NO) as released from human endothelial cells in vitro. The used approach is based on the following considerations a) use low concentrations of DAF-2 (0.1 micro M) in order to reduce the contribution of DAF-2 auto-fluorescence to the measured total fluorescence, and b) subtract the DAF-2 auto-fluorescence from the measured total fluorescence. The advantage of this method is the reliable quantification of NO in a biological system in the nanomolar range once thoroughly validated. Here we focus in addition to the previous publication (Leikert et al., FE... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591716 Wed, 01 Jan 2003 05:00:00 +0100 1591716 http://www.medworm.com/index.php?rid=1591715&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569612%26dopt%3DAbstract Authors: Malloff C, Dullaghan E, Li A, Stokes R, Fernandez R, Lam W We have developed two whole genome-scanning techniques to aid in the discovery of polymorphisms as well as horizontally acquired genes in prokaryotic organisms. First, two-dimensional bacterial genomic display (2DBGD) was developed using restriction enzyme fragmentation to separate genomic DNA based on size, and then employing denaturing gradient gel electrophoresis (DGGE) in the second dimension to exploit differences in sequence composition. This technique was used to generate high-resolution displays that enable the direct comparison of > 800 genomic fragments simultaneously and can be adapted for the high-throughput comparison of bacterial genomes. 2DBGDs are capable of detecting acquired and altered DNA, howeve... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591715 Wed, 01 Jan 2003 05:00:00 +0100 1591715 http://www.medworm.com/index.php?rid=1591714&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569613%26dopt%3DAbstract Authors: Maiato H, Sunkel CE, Earnshaw WC Here we describe a detailed methodology to study the function of genes whose products function during mitosis by dsRNA-mediated interference (RNAi) in cultured cells of Drosophila melanogaster. This procedure is particularly useful for the analysis of genes for which genetic mutations are not available or for the dissection of complicated phenotypes derived from the analysis of such mutants. With the advent of whole genome sequencing it is expected that RNAi-based screenings will be one method of choice for the identification and study of novel genes involved in particular cellular processes. In this paper we focused particularly on the procedures for the proper phenotypic analysis of cells after RNAi-mediated depletion of proteins required for... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591714 Wed, 01 Jan 2003 05:00:00 +0100 1591714 http://www.medworm.com/index.php?rid=1591713&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569614%26dopt%3DAbstract Authors: Jacobsson K, Rosander A, Bjerketorp J, Frykberg L Shotgun phage display cloning involves construction of libraries from randomly fragmented bacterial chromosomal DNA, cloned genes, or eukaryotic cDNAs, into a phagemid vector. The library obtained consists of phages expressing polypeptides corresponding to all genes encoded by the organism, or overlapping peptides derived from the cloned gene. From such a library, polypeptides with affinity for another molecule can be isolated by affinity selection, panning. The technique can be used to identify bacterial receptins and identification of their minimal binding domain, and but also to identify epitopes recognised by antibodies. In addition, after modification of the phagemid vector, the technique has also been used to identify bac... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591713 Wed, 01 Jan 2003 05:00:00 +0100 1591713 http://www.medworm.com/index.php?rid=1591712&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14569615%26dopt%3DAbstract Authors: Underkoffler LA, Collins JN, Choi JD, Oakey RJ Microsatellite markers are simple sequence repeats within the mammalian genome that can be used for identifying disease loci, mapping genes of interest as well as studying segregation patterns related to meiotic nondisjunction. Different strains of mice have variable CA repeat lengths and PCR based methods can be used to identify them, thus allowing for specific genotypes to be assigned. Molecular genotyping offers such identification at any developmental stage, which allows for a broad range of anomalies to be studied. We studied chromosomal segregation in relation to nondisjunction in early-gestation mouse embryos using molecular genotyping. Information on the parental origin as well as the number of chromosomes a given progeny ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591712 Wed, 01 Jan 2003 05:00:00 +0100 1591712 http://www.medworm.com/index.php?rid=1591711&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615812%26dopt%3DAbstract This report describes tools to study chromatin assembly in the model system Saccharomyces cerevisiae. Modifications to an in vitro chromatin assembly assay are described that allowed a brute force screen of temperature sensitive (ts) yeast strains in order to identify chromatin assembly defective extracts. This screen yielded mutations in genes encoding two ubiquitin protein ligases (E3s): RSP5, and a subunit of the Anaphase Promoting Complex (APC), APC5. Additional modifications are described that allow for a rapid analysis and an in vivo characterization of yeast chromatin assembly mutants, as well as any other mutant of interest. Our analysis suggests that the in vitro and invivo chromatin assembly assays are responsive to different cellular signals, including cell cycle cues that invol... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591711 Wed, 01 Jan 2003 05:00:00 +0100 1591711 http://www.medworm.com/index.php?rid=1591710&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615813%26dopt%3DAbstract Authors: Zhang X, Oglesbee M The 72 kDa heat shock protein (HSP72) is a molecular chaperone that binds native protein with low affinity. These interactions can alter function of the substrate, a property known as HSP-mediated activity control. In the present work, BIAcore instrumentation was used to monitor binding reactions between HSP72 and naturally occurring sequence variants of the measles virus (MV) nucleocapsid protein (N), a structural protein regulating transcription/replication of the viral genome. Binding reactions employed synthetic peptides mimicking a putative HSP72 binding motif of N. Sequences were identified that bound HSP72 with affinities comparable to well-characterized activity control reactions. These sequences, but not those binding with lesser affinity, supporte... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591710 Wed, 01 Jan 2003 05:00:00 +0100 1591710 http://www.medworm.com/index.php?rid=1591709&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615814%26dopt%3DAbstract Authors: Mahajan SD, Aalinkeel R, Schwartz SA, Chawda RP, Nair MP CD8+ cytotoxic T lymphocyte (CTL) activity is currently believed to be one of the key immunologic mechanisms responsible for the prevention or attenuation of HIV-1 infection. The induction of CD8+ T cell activation may also result in the production of soluble or non-classical lytic factors that are associated with protection from infection or slower disease progression. Traditionally, CD8+ CTL responses have been measured by the classic chromium release assay, monitoring the ability of T cells (Effector cells) to lyse radiolabelled HLA - matched "target cells" that express the appropriate antigen-MHC complex. This method is not only labor intensive, semi quantitative assay at best, but also needs fresh, non-cryopreserved... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591709 Wed, 01 Jan 2003 05:00:00 +0100 1591709 http://www.medworm.com/index.php?rid=1591708&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615815%26dopt%3DAbstract Authors: Kim W, Surette MG Salmonella enterica serovar Typhimurium is capable of swarming over semi-solid surfaces. Although its swarming behavior shares many readily observable similarities with other swarming bacteria, the phenomenon remains somewhat of an enigma in this bacterium since some attributes skew away from the better characterized systems. Swarming is quite distinct from the classic swimming motility, as there is a prerequisite for cells to first undergo a morphological transformation into swarmer cells. In some organisms, swarming is controlled by quorum sensing, and in others, swarming has been shown to be coupled to increased expression of important virulence factors. Swarming in serovar Typhimurium is coupled to elevated resistance to a wide variety of structurally and... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591708 Wed, 01 Jan 2003 05:00:00 +0100 1591708 http://www.medworm.com/index.php?rid=1591707&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615816%26dopt%3DAbstract Authors: Obermaier B, Dauer M, Herten J, Schad K, Endres S, Eigler A We developed a new 2-day protocol for the generation of dendritic cells (DCs) from human monocytes in vitro. First, we demonstrated that 24 hours of culture with GM-CSF and IL-4 are sufficient to generate immature DCs capable of antigen uptake. We then compared two different strategies for DC maturation: proinflammatory mediators were either added together with GM-CSF and IL-4 from the beginning of cell culture or added after 24 hours of differentiation with GM-CSF and IL-4. After 48 hours of total culture period, expression of activation markers was more pronounced in cells generated by the 2-step differentiation and activation method. Our new protocol for 2-day DC differentiation reduces labor, cost and time and als... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591707 Wed, 01 Jan 2003 05:00:00 +0100 1591707 http://www.medworm.com/index.php?rid=1591706&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615817%26dopt%3DAbstract Authors: Azam M, Raz T, Nardi V, Opitz SL, Daley GQ The discovery of oncogenes and signal transduction pathways important for mitogenesis has triggered the development of target-specific small molecule anti-cancer compounds. As exemplified by imatinib (Gleevec), a specific inhibitor of the Chronic Myeloid Leukemia (CML)-associated Bcr-Abl kinase, these agents promise impressive activity in clinical trials, with low levels of clinical toxicity. However, such therapy is susceptible to the emergence of drug resistance due to amino acid substitutions in the target protein. Defining the spectrum of such mutations is important for patient monitoring and the design of next-generation inhibitors. Using imatinib and BCR/ABL as a paradigm for a drug-target pair, we recently reported a retroviral... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591706 Wed, 01 Jan 2003 05:00:00 +0100 1591706 http://www.medworm.com/index.php?rid=1591705&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14615818%26dopt%3DAbstract This report summarizes the methods we used in establishing this murine model. By treating murine CD4+ T cells with DNA hypomethylating agents and by transfection we were able to test the in vitro effects of integrin overexpression on T cell autoreactive proliferation, cytotoxicity, adhesion and trafficking. Furthermore, we showed that the ability to induce in vivo autoimmunity may be unique to the integrin lymphocyte function associated antigen-1 (LFA-1). PMID: 14615818 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591705 Wed, 01 Jan 2003 05:00:00 +0100 1591705 http://www.medworm.com/index.php?rid=1591704&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14647536%26dopt%3DAbstract Authors: Sirchia R, Ciacciofera V, Luparello C It is widely acknowledged that the presence of extracellular matrix components as substrates can drastically modulate the phenotype and gene expression of cultured cells, including tumor cells. A number of published reports indicated that substrates made from two peculiar collagen species, i.e. type V and OF/LB, which are abnormally deposited in the stroma of primary ductal infiltrating carcinoma (d.i.c.) of the breast "in vivo," were able to exert marked and opposite effects on "in vitro" viability, growth and invasiveness of the 8701-BC cell line, isolated from d.i.c.-affected breast epithelium. To complement such functional data on the effect of cell-collagen interactions with information at molecular level, we have utilized a combinati... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591704 Wed, 01 Jan 2003 05:00:00 +0100 1591704 http://www.medworm.com/index.php?rid=1591703&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D14702098%26dopt%3DAbstract Authors: Pridgeon JW, Geetha T, Wooten MW The UBA domain is a conserved sequence motif among polyubiquitin binding proteins. For the first time, we demonstrate a systematic, high throughput approach to identification of UBA domain-interacting proteins from a proteome-wide perspective. Using the rabbit reticulocyte lysate in vitro expression cloning system, we have successfully identified eleven proteins that interact with p62's UBA domain, and the majority of the eleven proteins are associated with neurodegenerative disorders, such as Alzheimer's disease. Therefore, p62 may play a novel regulatory role through its UBA domain. Our approach provides an easy route to the characterization of UBA domain interacting proteins and its application will unfold the important roles that the UBA do... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591703 Wed, 01 Jan 2003 05:00:00 +0100 1591703 http://www.medworm.com/index.php?rid=1591723&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734596%26dopt%3DAbstract Authors: Sklodowaka A, Wozniak M, Matlakowska R In this paper, we present our method for the measurement of contact angles on the surface of minerals during the bioleaching process because the standard deviation obtained in our measurements achieved unexpectedly low error. Construction of a goniometer connected with a specially prepared computer program allowed us to repeat measurements several times over a short time course, yielding excellent results.After defining points on the outline of the image of a drop and its baseline as well of the first approximation of the outline of the drop, an iterative process is initiated that is aimed at fitting the model of the drop and baseline. In turn, after defining the medium for which measurements were made, the work of adhesion is determined ... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591723 Thu, 29 Apr 1999 04:00:00 +0100 1591723 http://www.medworm.com/index.php?rid=1591722&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734597%26dopt%3DAbstract Authors: Ramalho-Santos J, Pedroso De Lima MC It had been suggested that influenza virus-mediated membrane fusion might be dependent on a pH gradient across a target membrane. We have designed experiments in which this issue could be addressed. Two populations of liposomes were prepared, both simulating the plasma membrane of target cells, but with the pH of the internal aqueous medium buffered either at pH 7.4 (physiological cytosol pH) or at pH 5.0 (endosomal pH at which influenza virus displays maximal fusion activity). By monitoring fusion using the R18 assay, we found that the internal pH of the target liposomes did not influence membrane merging as mediated by the influenza virus hemagglutinin, thus demonstrating that a transmembrane pH gradient is not required in this fusion pro... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591722 Tue, 16 Mar 1999 05:00:00 +0100 1591722 http://www.medworm.com/index.php?rid=1591724&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734595%26dopt%3DAbstract This article attempts to convey the joys and frustrations of skimming the Internet trying to find relevant information concerning an academic's work as a scientist, a student or an instructor. A brief overview of the Internet and the "do's and don'ts" for the neophyte as well for the more seasoned "navigator" are given. Some guidelines of "what works and what does not" and "what is out there" are provided for the scientist with specific emphasis for biologists, as well as for all others having an interest in science but with little interest in spending countless hours "surfing the net". An extensive but not exhaustive list of related websites is provided. PMID: 12734595 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591724 Fri, 08 Jan 1999 05:00:00 +0100 1591724 http://www.medworm.com/index.php?rid=1591721&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734598%26dopt%3DAbstract Authors: Marín D, Pérez P, Teijeiro C, Palecek E Stripping voltammetry (SV) is used to quantitatively determine concentrations of the anti-neoplastic drug mitomycin C (MMC) alone and in mixtures with 5-fluorouracil and cisplatin, both of which are used in combined chemotherapy with MMC. If the accumulation is performed at the potentials of MMC reduction (-0.35 V vs. SCE), reduced MMC is strongly adsorbed at the electrode. It is possible to prepare a MMC-modified electrode, which, after a washing step, is transferred to the background electrolyte to determine MMC by voltammetry. This procedure, which is termed transfer stripping voltammetry (TSV), helps to eliminate interferences and can be applied for a direct determination of MMC alone or in mixtures with other drugs in urin... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591721 Thu, 17 Sep 1998 04:00:00 +0100 1591721 http://www.medworm.com/index.php?rid=1591720&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734599%26dopt%3DAbstract Authors: Lee JI, Woo SK, Kim KI, Park KC, Baek SH, Yoo YJ, Chung CH A general method for the assay of deubiquitinating enzymes was described in detail using (125)I-labeled ubiquitin-fused alphaNH-MHISPPEPESEEEEEHYC (referred to as Ub-PESTc) as a substrate. Since the tyrosine residue in the PESTc portion of the fusion protein was almost exclusively radioiodinated under a mild labeling condition, such as using IODO-BEADS, the enzymes could be assayed directly by simple measurement of the radioactivity released into acid soluble products. Using this assay protocol, we could purify six deubiquitinating enzymes from chick skeletal muscle and yeast and compare their specific activities. Since the extracts of E. coli showed little or no activity against the substrate, the assay protocol shoul... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591720 Mon, 20 Jul 1998 04:00:00 +0100 1591720 http://www.medworm.com/index.php?rid=1591730&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734589%26dopt%3DAbstract Authors: Konno R Four methods (an enzyme activity assay, western blotting, RT-PCR, and northern hybridization) to detect the enzyme D-amino-acid oxidase are described. PMID: 12734589 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591730 Thu, 14 May 1998 04:00:00 +0100 1591730 http://www.medworm.com/index.php?rid=1591729&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734590%26dopt%3DAbstract Authors: Yeates C, Gillings MR, Davison AD, Altavilla N, Veal DA Amplification of DNA from soil is often inhibited by co-purified contaminants. A rapid, inexpensive, large-scale DNA extraction method involving minimal purification has been developed that is applicable to various soil types (1). DNA is also suitable for PCR amplification using various DNA targets. DNA was extracted from 100g of soil using direct lysis with glass beads and SDS followed by potassium acetate precipitation, polyethylene glycol precipitation, phenol extraction and isopropanol precipitation. This method was compared to other DNA extraction methods with regard to DNA purity and size. PMID: 12734590 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591729 Thu, 14 May 1998 04:00:00 +0100 1591729 http://www.medworm.com/index.php?rid=1591728&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734591%26dopt%3DAbstract This study showed the benefits of employing several different complementary methods to investigate bacterial enzyme activity. PMID: 12734591 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591728 Thu, 14 May 1998 04:00:00 +0100 1591728 http://www.medworm.com/index.php?rid=1591727&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734592%26dopt%3DAbstract Authors: Becerra M, Cerdãn E, Gonzãlez Siso MI Several micro-scale chromatography-based procedures for purification of the beta-galactosidase from the yeast Kluyveromyces lactis were assayed. Purified enzyme was suitable to be used as antigen to induce polyclonal antibodies production. Specific staining of non-denaturing PAGE gels with chromogenic substrates allowed the determination of the number of subunits forming the native enzyme. PMID: 12734592 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591727 Thu, 14 May 1998 04:00:00 +0100 1591727 http://www.medworm.com/index.php?rid=1591726&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734593%26dopt%3DAbstract Authors: Deng HW Deng and Lynch (1, 2) proposed to characterize deleterious genomic mutations from changes in the mean and genetic variance of fitness traits upon selfing in outcrossing populations. Such observations can be readily acquired in cyclical parthenogens. Selfing and life-table experiments were performed for two such Daphnia populations. A significant inbreeding depression and an increase of genetic variance for all traits analyzed were observed. Deng and Lynch's (2) procedures were employed to estimate the genomic mutation rate (U), mean dominance coefficient (), mean selection coefficient (), and scaled genomic mutational variance (). On average, and (^ indicates an estimate) are 0.84, 0.30, 0.14 and 4.6E-4 respectively. For the true values, the and are lower bounds, and a... Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591726 Thu, 14 May 1998 04:00:00 +0100 1591726 http://www.medworm.com/index.php?rid=1591725&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734594%26dopt%3DAbstract Authors: Bonora GM, Zaramella S, Veronese FM The chemical modification of synthetic oligonucleotides has recently been investigated to improve their pharmacological utilization. In addition to chemical alterations of the backbone and of the heterocyclic bases, their conjugation with amphiphylic moieties, such as the polyethylene glycol has been proposed. The large scale production of these molecules as demanded for commercial purposes is hampered by the heterogeneity of the solid-phase processes and by the low reactivity of high-molecular weight PEGs in solution. A new synthetic procedure based on the recently developed liquid-phase method (HELP), has been set up to overcome these limitations. PMID: 12734594 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591725 Thu, 14 May 1998 04:00:00 +0100 1591725 http://www.medworm.com/index.php?rid=1591719&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734600%26dopt%3DAbstract This study demonstrates the feasibility of such experiments with PLBs, amenable to physical constraints, and thus offers new opportunities for systematic studies of structure-function relationships in membrane-associating molecules. PMID: 12734600 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591719 Thu, 14 May 1998 04:00:00 +0100 1591719 http://www.medworm.com/index.php?rid=1591718&cid=s_37597_67_f&fid=37597&url=http%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fentrez%2Fquery.fcgi%3Ftmpl%3DNoSidebarfile%26db%3DPubMed%26cmd%3DRetrieve%26list_uids%3D12734601%26dopt%3DAbstract Authors: Breierova E Six extracellular yeast glycoproteins were prepared from three yeast species in osmotic equilibrium and unequilibrium environments and used as non-penetrating cryoadditives. Glycoproteins secreted by the strain Dipodascus australiensis into growth medium containing NaCl (8% w/v) were found to be the most effective cryoadditives. It was possible to use these glycoproteins alone (without DMSO as penetrating agent) for the cryoprotection of the studied yeasts. PMID: 12734601 [PubMed - as supplied by publisher] (Source: Biological Procedures Online) Biological Procedures Online journals http://www.medworm.com/rss/comments.php?id=1591718 Thu, 14 May 1998 04:00:00 +0100 1591718